White adipose tissue-infiltrated CD11b+ myeloid cells are a source of S100A4, a new potential marker of hepatic damage

in European Journal of Endocrinology
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  • 1 Department of Endocrinology and Nutrition, Germans Trias i Pujol Research Institute
  • 2 Department of Biochemistry and Physiology, School of Pharmacy and Food Sciences, Institut de Biomedicina de la Universitat de Barcelona (IBUB), Universitat de Barcelona, Barcelona, Spain
  • 3 Department of Biochemistry and Molecular Biology, and Institute of Biomedicine, University of Barcelona, Barcelona, Spain
  • 4 Biomedical Research Center (Red Fisiopatología de la Obesidad y Nutrición) (CIBEROBN), ISCIII, Madrid, Spain
  • 5 Research Institute and Infectious Diseases Unit, Hospital de la Santa Creu i Sant Pau, Barcelona, Spain
  • 6 Biomedical Research Center (Red Fisiopatología de la Diabetes y enfermedades metabólicas) (CIBERDEM), ISCIII, Madrid, Spain
  • 7 Department of Surgery, Germans Trias i Pujol Research Institute, Barcelona, Spain
  • 8 Department of plastic Surgery, Germans Trias i Pujol Research Institute, Barcelona, Spain
  • 9 Área de Bioquímica y Biología Molecular, Departamento de Ciencias Básicas de la Salud, Facultad de Ciencias de la Salud, Universidad Rey Juan Carlos, Avda. de Atenas s/n. Alcorcón, Madrid, Spain

Correspondence should be addressed to L Herrero or D Sánchez-Infantes; Email: lherrero@ub.edu or dsanchez@igtp.cat

*(M Reyes, L González and K Ibeas contributed equally to this work)

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Context

The endocrine and immunological properties of subcutaneous vs visceral adipose tissue (sWAT and vWAT, respectively) have turned a milestone in the study of metabolic diseases. The cytokine S100A4 is increased in obesity and has a role in adipose tissue dysfunction. However, the cellular source and its potential role in hepatic damage in obesity has not been elucidated.

Objective

We aim to study the regulation of S100A4 in immune cells present in sWAT and vWAT, as well as its potential role as a circulating marker of hepatic inflammation and steatosis.

Design

A cohort of 60 patients with obesity and distinct metabolic status was analyzed. CD11b+ myeloid cells and T cells were isolated from sWAT and vWAT by magnetic-activating cell sorting, and RNA was obtained. S100A4 gene expression was measured, and correlation analysis with clinical data was performed. Liver biopsies were obtained from 20 patients, and S100A4 circulating levels were measured to check the link with hepatic inflammation and steatosis.

Results

S100A4 gene expression was strongly upregulated in sWAT- vs vWAT-infiltrated CD11b+ cells, but this modulation was not observed in T cells. S100A4 mRNA levels from sWAT (and not from vWAT) CD11b+ cells positively correlated with glycemia, triglycerides, TNF-α gene expression and proliferation markers. Finally, circulating S100A4 directly correlated with liver steatosis and hepatic inflammatory markers.

Conclusion

Our data suggest that sWAT-infiltrated CD11b+ cells could be a major source of S100A4 in obesity. Moreover, our correlations identify circulating S100A4 as a potential novel biomarker of hepatic damage and steatosis.

Supplementary Materials

    • Supplementary Table S1. Primers pairs for qRT-PCR detection and relative quantification of genes in humans.
    • Supplementary Table S2. Correlations between the S100A4 relative gene level of sWAT and vWAT CD11b+ cells and clinical parameters.
    • Supplementary Figure S1. mRNA expression levels of S100A4 in T cells and CD11b+ cells isolated from sWAT and vWAT, and percentage of CD11b+ cells isolated from both tissues. S100A4 mRNA expression levels in T cells vs. CD11b+ cells present in sWAT (A) and vWAT (B). C) % CD11b+ cells that were isolated from sWAT and vWAT from the above described cohort of patients with obesity. D) Purity of CD11b+ and CD3+ measured by citometer. Lines indicate the mean values for each group. Each dot represents a single individual. ns, P > 0.05. *, P ≤ 0.05. **, P ≤ 0.01. ***, P ≤ 0.001. ****, P ≤ 0.0001.
    • Supplementary Figure S2. S100A4 protein levels in stromo-vascular fraction (SVF) and adipocytes. The protein levels were obtained from the SVF, adipocytes and the whole adipose tissue, and measured by ELISA. S100A4 protein levels in SVF compared to adipocytes. ns, P > 0.05. *, P ≤ 0.05. **, P ≤ 0.01.
    • Supplementary Figure S3. Correlation between S100A4 gene expression of human adipose tissue resident CD11b+ cells and markers of inflammation and quiescence. Plots showing the linear correlation between S100A4 gene expression of sWAT resident CD11b+ cells with (A) IFNgamma gene expression, (B) IL-1ß gene expression, (C) CD86 gene expression, (D) GADD45G gene expression, (E) ITGB1 gene expression, (F) GLS2 gene expression.
    • Supplementary Figure S4. S100A4 gene expression is not modulated in M1 and M2 in vitro macrophages. Total RNA was extracted from RAW 264.7 macrophages cell line pretreated with 60-ng/mL LPS or 40-ng/mL IL4 respectively for 12 hours and the controls. The mRNA expression of S100A4 was quantified using RT-qPCR. (A) TNFα gene expression in control, M1 and M2 macrophages, (B) ARG1 gene expression in control, M1 and M2 macrophages, and (C) S100A4 gene expression in control, M1 and M2 macrophages.
    • Supplementary Figure S5. S100A4 gene expression in adipose tissue vs liver from patients with obesity. Total RNA was extracted from vWAT and liver from patients that underwent bariatric surgery. The mRNA expression of S100A4 was quantified using RT-qPCR.

 

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